PROJECTSUMMARY ALT (Alternative Lengthening of Telomeres) cancers encompass a significant fraction (15%) of human cancers. These immortalized cells maintain their telomeres using homology-directed recombination. The observation that ALT is activated in a significant fraction of tumors, coupled with its potential as an adaptive mechanism to anti-telomerase tumor therapies, make it an important target for anti-cancer strategies. Yet, targetedtherapeuticsforALTcancershavenotbeengreatlyexplored.Recentstudiesshowedthatahallmark of ALT cells is loss of the chromatin remodeler ATRX, but how its loss contributes to telomere recombination had not been elucidated. We found that loss of ATRX suppresses resolution of sister chromatid cohesion at telomeres to promote sister telomere recombination and prevent deleterious non-allelic recombination. Sister telomere cohesion is normally resolved in G2/prophase of the cell cycle by the poly-ADP ribose polymerase tankyrase1thatlocalizestotelomeresthroughtheshelterinsubunitTRF1.WefoundthatduetolossofATRX the?normal?systemoftelomereresolutionisdefectiveinALTcellsandtelomeresremaincoheredintomitosis. IntheabsenceofATRXitsbindingpartner(thehistonevariantmacroH2A1.1)isfreetosequestertankyrase1, preventingit fromresolvingtelomerecohesion.Persistenttelomerecohesionhasanessentialrole,asforced resolutionoftelomerecohesionbyATRXintroductionortankyrase1overexpressioninALTcellsleadstoloss ofrecombinationbetweensistertelomeres,increasednon-sistercopying,andcelldeath.Wehypothesizethat persistenttelomerecohesionassistsintelomeremaintenancebypromotingsistertelomererecombinationand in safeguarding genome integrity by preventing aberrant and toxic non-sister recombination. In Aim 1 of this proposal, I will elucidate the mechanisms of persistent telomere cohesion and in Aim 2, I will define the proteome of the cohered state in ALT cells. My studies will provide mechanistic insights into telomere recombinationandidentifypotentialtargetsforALTcancertherapy.